1. Technical Field
The present disclosure relates to a nucleic acid sequence measuring method, a nucleic acid sequence measuring device, a manufacturing method for nucleic acid sequence measuring device, and a nucleic acid sequence measuring apparatus.
2. Background Art
A nucleic acid sequence measuring method has been known in which a target having a particular nucleic acid sequence in a sample is measured through hybridization between the target and a probe by using a nucleic acid sequence measuring device. For example, a method of checking the presence or absence of, or measuring the amount of a predetermined nucleic acid by using a DNA chip as a nucleic acid sequence measuring device has been widely known. The DNA chip includes a detection probe with a sequence complementary with the nucleic acid to be detected, the detection probe being fixed to the solid surface of a substrate or the like. The detection probe is fixed to the solid surface at any of 3′ end and 5′ end thereof. On the other hand, the nucleic acid to be detected (target) is modified with a fluorescent molecule or the like.
For example, the measurement using the DNA chip is conducted based on procedures as below.
(1) The detection target nucleic acid molecule in the sample is amplified by a nucleic acid amplifying technique such as PCR. Moreover, a fluorescent molecule is added to the amplified molecule (i.e., the nucleic acid to be detected, or target). The fluorescent molecule is added in a manner that, for example, the nucleic acid with the fluorescent pigment added thereto is mixed in the PCR amplification or a primer to which the fluorescent pigment is added in advance is used in the PCR amplification. Alternatively, the fluorescent pigment is added by chemical modification after the amplification.
(2) A solution containing the target is prepared and added to the DNA chip having the detection probe. The target modified with the fluorescence is captured by the detection probe through the hybridization with the detection probe.
(3) By washing the DNA chip, the fluorescence emitted from the molecules that are not captured or the molecules that are coupled nonspecifically to the nucleic acid sequence of the detection probe is eliminated. This washing step is repeated for a plurality of times depending on the desired degree of washing. After the DNA chip is washed and before the DNA chip is measured, the solid surface is dried up.
(4) The solid surface is observed using a fluorescence reading apparatus. The presence or absence of the target in the sample is checked based on whether the detection probe of the DNA chip exhibits the fluorescence.